1252737 玖、發明說明 (發明說明應敘明：發明所屬之技術領域、先前技術、內容、實施方式及圖式簡單說明) (一） 發明所屬之技術領域： 本發明係關於一種改良的醬油釀造方法，其包括將醬油 麴菌混入一種鹽水中以製成醬油醪，將預培養之醬油乳酸 菌及醬油酵母菌添加至醬油醪中，在以一般技術添加之後 進行醬油醪的醱酵及熟化，以及關於一種可確實獲得具有 所需乙醇濃度及乳酸濃度和具有甘醇風味醬油的方法。 (二） 先前技術 傳統的醬油釀造法係包括將蒸過的黃豆與烤過及碾製過 的小麥摻混在一起，並以用於醬油的種子麴菌接種於此混 合物，並且培養這種混合物以製得醬油麹菌，在醬油麴菌 中添加適量的鹽水以獲得一種醬油醪，使醬油醪經過一段 固定時間的醱酵及熟化之後，獲得一種熟化的醬油醪，最 後再將這種熟化的醬油醪予以壓濾。然而，醬油醪的醱酵 和熟化通常是在開放的系統中進行，並且幾乎沒有人工添 加任何微生物，如醬油酵母菌或醬油乳酸菌。因此，所有 在醬油_中作用的微生物（酵母菌和乳酸菌）皆是來自棲息 於釀造地點、設施、裝置等之天然植物所衍生的天然雜質 〇 在本發明中’所謂的，，野生酵母菌，，係指棲息於釀造醬油 之地點、設施、裝置等處’並且自然會在醬油醪中污染和 ~ 6 - 1252737 增殖之酵母菌。 相反地’本發明中所使用的醬油酵母菌很容易藉由已知 的一般方式自醬油醪中收集（分離），並且以能夠產生酒精 且能耐鹽者爲較佳。這種醬油酵母菌純粹是用人工培養， 並且被添加至醬油醪中。因此，這種經過預培養的醬油酵 母菌顯然與野生酵母菌有很大的不同。 天然植物是相當多樣化的，因此，天然污染的醬油酵母 菌及醬油乳酸菌中可能會含有具有不是完全有利用醬油品 質之特性的細菌。此外，由於天然植物因地區或時間的不 同而會有不同組成差異的此項缺點，因於產品的品質會隨 著釀造地點的不同而有所改變，而無法在一整年內獲得具 有一定品質的醬油。 基於這些觀點’爲了能連續製造出具有一定品質之釀造 醬油而無須考慮地區或時間等限制，本發明提出了 一種釀 造醬油的方法。這種釀造醬油的方法包括將醬油麴菌混入 一種鹽水中以製成醬油醪；將預培養之醬油乳酸菌及醬油 酵母菌添加至醬油醪中；然後在一般技術添加之後進行醬 油醪的醱酵及熟化。 說的更明確一點，一種已知的方法，包括將醬油麴菌混 入一種鹽水中以製成醬油醪，將預培養之醬油乳酸菌添加 至醬油醪中，培養這些細菌，在進行乳酸醱酵的時候（通常 爲混合後的20到40天），當醬油醪中之乳酸菌細胞數超過 1 〇7個/每克醬油醪且醬油醪的pH値降低至5 . 1到5 . 2之間 時’加入醬油酵母菌（主要爲 Zygosaccharomyces rouxii 1252737 ’它是主要的醬油醱酵酵母菌），進行乙醇醱酵，因而製得 了具有良好風味的釀造醬油（醬油的科學與技術，由 Shinrokuro TOCHIKURA所編輯及撰寫，日本釀造學會， 1 9 8 8 )(食品微生物學手冊，和井久雄，金子安之及山口和 夫編撰，技報堂出版，1 9 9 5 )。 然而在此方法中，由於使用的不是無菌的麴，並且是在 微生物的開放容器中進行醱酵及熟化，基於複雜的麻煩因 素無法被界定，因此醬油醪的乳酸醱酵及酵母醱酵都很容 易受到野生微生物的污染。因此，很難在相當長的時段內 維持适些被添加微生物得以穩定且平衡的生長，同時又要 能兼顧較佳的醱酵效果。 此外，在純培養的情況下，雖然主要的醬油醱酵酵母菌 種具有高的乙醇生產力或者是能夠形成良好的醬油氣味， 但是當這些菌種被用於真正的釀造製程中時，在許多情況 下並無法達到純培養條件下的相同結果。 因此，近幾年來，有一些硏究企圖能使得添加之醬油酵 母菌和乳酸菌達成平衡且穩定的長期生長。說的更明確一 點，即使在最新的、密封及獨立型混合及醱酵槽中，以及 鄰近的混合及醱酵槽之間控制進醬油醪的醱酵和熟化，並 且在每一次使用之後，就將混合及醱酵槽或是混合及醱酵 的路徑予以淸潔。 然而，因爲不同批次之間的醬油麴菌或醬油醪的品質亦 有所不同，使得pH値會隨著乳酸醱酵作用而呈現不穩定下 降，使得這種方法也有缺點，因此，只能依照專家的經驗 -8- 1252737 和感覺來判斷添加酵母菌的適當時機。 此外’要自淸潔的混合及醱酵槽或混合及醱酵路徑中完 全淸除微生物是有其極限的。因此，當耐鹽性的野生酵母 菌無法被消滅時，會有少數的野生酵母菌殘留並且污染了 醬油醪’並且可能會在添加醬油酵母菌之前就成長成優勢 的一部分，不但會抑制乳酸醱酵，同時也會受到野生酵母 菌的影響而產生不良的氣味。 此外，被添加的酵母菌須先被培養成所需的量並且予以 貯存。有一項缺點是：當酵母菌的添加時間明顯地與預定 時間要早或晚時，可能會導致添加酵母菌不足或是酵母菌 活性降低等結果’因此酵母菌的醱酵作用並不是在添加至 醬油醪之後立即開始的。 (三）發明內容 本發明的目的之一係可靠的獲得具有所需乙醇濃度（例如 3 % ( V / V )或更高）、所需乳酸濃度（例如〇 . 8到1 . 〇 % )、ρ Η値 爲4 . 6到4 . 9之間且具有甘醇風味的醬油，其係藉由一種 醬油的釀造方法來解決上述的缺點，其方法包括將醬油麴 菌混入一種鹽水中以製成醬油醪，將預培養之醬油乳酸菌 及醬油酵母菌添加至醬油醪中，在以一般技術添加之後， 進行醬油醪的醱酵及熟化。 將硏究的重點放在解決上述問題之結果，本發明人發現 :藉由注意醬油醪中耐鹽性野生酵母菌在醱酵初期的細胞 數，並且在醬油醪中添加比野生酵母菌多上1 〇到丨〇〇倍的 醬油酵母活菌，可達到抑制野生酵母菌成長的效果，同時 -9- 1252737 也不會影響乳酸醱酵，只會活化添加之醬油酵母菌的成長 ，並且可達到成功的乙醇醱酵作用。此外，本發明人亦發 現：當醬油酵母菌係在混合之後的1 0天內添加至醬油醪中 ，而使得醬油酵母活菌的細胞數比既存於醬油麴中的野生 酵母菌多1 0倍到1 0 0倍時，僅添加極少量的醬油酵母菌即 足以達成上述的效果。此外，本發明者發現：在此期間， 如果醬油酵母菌的添加量超過2 X 1 0 5個細胞/每克醬油酸 時’共存於醬油醪中的乳酸菌所進行的醱酵作用就會受到 影響’而使得乳酸不會被生成和累積，而導致醬油的風味 不佳。此外，本發明者亦發現；如果醬油酵母菌以2 X 1 〇5 或更少個細胞/每克醬油醪的比率添加，並使得醬油酵母菌 的細胞數比既存於醬油醪中的野生酵母菌多1 〇倍到1 〇〇倍 曰寸’醬油酵母菌所進行的乙醇醱酵可充份進行，而不會抑 制A酸醱酵’並且可以可靠的獲得具有所需乙醇濃度（例如 3%( V/V)或更高）、所需乳酸濃度（例如〇 . 8到〇%)、pH値 爲4 . 6到4 . 9之間且具有甘醇風味的醬油。 本發明已藉由這些發現而得以完成。說的更明確一點， 本發明係關於一種醬油的釀造方法，其包括步驟爲：將醬 油麴菌混入一種鹽水中以製成醬油醪；將預培養之醬油乳 酸菌及醬油酵母菌添加至醬油醪中；在以一般技術添加之 後，進行醬油醪的醱酵及熟化，其中，被添加至醬油醪中 的k油酵母菌爲醬油酵母菌的細胞數比已存在於醬油醪中 之野生酵母菌大上至1〇〇倍。 本發明係一種醬油的釀造方法，包括步驟爲：將醬油麴 -10- 1252737 菌混入一種鹽水中以製成醬油醪；將預培養之醬油乳酸菌 及醬油酵母菌添加至醬油醪中；並且在以一般技術添加之 後，進行醬油醪的醱酵及熟化，其中被添加至醬油醪中的 醬油酵母菌係在混合1 〇天內添加，以醬油酵母菌細胞數比 已存在於醬油醪中之野生酵母菌多上1 0至1 0 0倍之量添加 〇 此外，本發明係一種醬油的釀造方法，包括步驟爲：將 醬油麹菌混入一種鹽水中以製成醬油醪；將預培養之醬油 乳酸菌及醬油酵母菌添加至醬油醪中；並且在以一般技術 添加之後，進行醬油醪的醱酵及熟化，其中被添加至醬油 醪中的醬油酵母菌係在混合1 0天內添加，以醬油酵母菌細 胞數比已存在於醬油醪中之野生酵母菌多上10至100倍之 量添加，並且爲2 X 1 0 5或更少個細胞/每克醬油醪。 本專利申請書包含日本專利申請書第200 1 - 3 8 8 748號中 所揭示之部分或全部內容和/或圖示，該申請書爲本申請案 的優先權文件。 以下將針對本發明做更詳細的描敘。 爲了實現本發明，首先，先將依一般生產醬油麴菌方法 所製備的醬油麴菌依照一般的醬油釀造方法混入鹽水中。 本文中所使用鹽水的用量比醬油麴菌原料的重量多1到 3倍。這裡所使用的食鹽濃度係經調整成適合釀造醬油所 用，使食鹽的濃度爲1 5到2 0 % ( W / V )，又以1 6到1 8 % ( W / V ) 爲更佳。在混合時，將食鹽的濃度調整在上述範圍內是相 S重女的，因爲食鹽、丨辰度低於1 5 %時，可能會使得醬油酸 1252737 在夏季25到35T：的溫度下腐敗。醬油醪的食鹽濃度較低 較利於酵素反應成功的進行，亦有利於醬油乳酸菌和醬油 酵母菌的增生及醱酵作用更爲順利，而使得醬油醪的細胞 溶fe及成熟更爲快速。然而’在食鹽濃度低於1 5 %的情況 下’所謂的腐敗細菌會大量成長’因而產生明顯酸的氣味 和酸敗味，終而使得醬油產品完全失去價値。因爲食鹽濃 度不當所造成的醬油醪腐敗可以藉由設定條件的方式來解 決’ s者如將醬油醪的溫度維持在4 0 °C或更高，或者是藉由 添加酸，例如鹽酸或乳酸的方式將pH値調整成3 . 〇或更低 。然而，這些條件同時也會破壞醬油的原有風味，因而使 得產品失去醬油的價値。當醬油醪的溫度或ρί]値或者是醬 油醪液的氮濃度不適合腐敗細菌的增殖或生存時，即使食 鹽的濃度低，醬油醪也不會腐敗。然而，這些只是特殊情 形，這些不能被視爲釀造醬油的一般方法。除非在夏季食 鹽的濃度爲17%或更高’或者是在春或秋季食鹽的濃度爲 1 5 %或更高，否則無法達到成功的醱酵和熟化作用。除此之 外’在4 0 °C或更高的溫度下水解醬油醪也會導致有不佳氣 味伴隨著醬油的缺點，這就是所謂的“ warm_brew，，（暖釀） 氣味。 接下來’將預先選好或特殊培育之具有良好性質的醬油 乳酸菌及醬油酵母菌（在下文中，有時被簡稱爲，，主要醬油 醱酵酵母菌”）添加至所製成的醬油醪中，除此之外，可依 照傳統釀造醬油的方法來添加醬油乳酸菌，其比率爲1 X丨〇4 個細胞/每克醬油醪到1 X 1 07個細胞/每克醬油醪之間。 - 1 2 - 1252737 在本發明中相當重要的一點是，所添加的醬油酵母菌可 使得醬油酵母菌的細胞數比已存在於醬油醪中之野生酵母 菌多上1 0至1 0 0倍。當此比率低於]0倍時會產生一種危 險’使得共存於醬油醪中的野生酵母菌變爲相對多數，而 具有良好性質且已被添加至醬油醪中的醬油酵母菌種就很 難大重成長’迨是由於其與共存於醬油酸中之野生酵母菌 的競爭之故，因此在競爭性的生長抑制作用之下，這些醬 油酵母菌可能就被消滅或是驅逐了。 相反地’如果添加的比率大於1 0 0倍時，共存的乳酸菌 不能活潑的成長’因此不能獲得具有所需乳酸濃度、適當 Ρ Η値（例如ρ Η値爲4 . 6到4 · 9之間）和甘醇風味的釀造醬 油。 此外’預培養的醬油乳酸菌和醬油酵母菌較好是在混合 後1 〇天以內添加’又以在混合後七天之內添加爲宜。最佳 的添加時間是在混合當時（剛混合之後）到混合後的隔天之 間。這個範圍會較爲適當的原因是因爲它可以大幅節省所 添加醬油酵母菌的用量。 相反地’當醬油酵母菌的添加時間是在混合後的丨丨天或 更晚時’共存在醬油醪中的野生酵母菌數變得更多。在這 @情況下，必需添加較大量的醬油酵母菌。結果造成，當 _母菌的濃度變的愈高時，就抑制了乳酸菌的生長，因此 ’這樣的結果將會造成所獲得的醬油具有低的乳酸含量且 風^未不佳。此外，還有一種危險是：被添加至醬油醪中的 胃 '油酵母菌（具有良好性質的主要醬油醱酵酵母菌）很難活 -13- 1252737 潑的成長’這是由於其與共存於醬油醪中之野生酵母菌的 競爭之故’因此在野生酵母菌的競爭性生長抑制作用之下 ’這些醬油酵母菌可能就被消滅或是驅逐了。也就是說， 因爲醬油酵母菌可能無法有效的增殖，而造成本發明的問 題無法順利被解決的危險。 相當重要的是，所添加醬油酵母菌的用量比率爲2 X 1 05 或更少個細胞/每克醬油醪。當用量的比率爲2 X 1 0 5個細 胞/每克醬油醪或更高時，乳酸菌將無法有效的增値。因此 ’可能無法獲得具有所需乳酸濃度、適當p pj値（例如ρ η値 爲4 . 6到4 . 9之間）和甘醇風味的醬油。 Μ本發明之醬油酵母菌的實例包括釀造醬油用的耐鹽 丨生醬油酵母囷’如：屬於Z y g 〇 s a c c h a r 〇 m y c e s r 〇 u X i i菌種 ’且主要適用於醬油醪之乙醇醱酵作用的主要醬油醱酵酵 母菌’其在6到28%(W/V)的氯化鈉和葡萄糖存在的情況下 ’具：有相當明顯的乙醇醱酵能力；屬於假絲酵母（Candicia) 屬’包括 Candida versatil is 和 Candida etch ellsii 的 耐鹽丨生酵母囷群；或者是T o r u 1 o p s i s屬的耐鹽性酵母菌群 ’其係當醬油主要醱酵酵母菌的乙醇醱酵完全之後才開始 作用’主要係關於醬油一般香氣的產生，有時亦被稱爲後 熟酵母菌群。 在追些酵母菌群中’具有活性乙醇醱酵能力的主要醬油 釀酵酵母囷以可獲得（特別是）具有3 % ( V / V )或更高乙醇濃 度和甘醇風味的爲較佳。 依照本發明人的硏究’主要的醬油醱酵酵母菌不能被視 -14- 1252737 爲共同的，因爲它們會隨著菌種的不同而有所差異。也就 是說，不同菌種的主要醬油醱酵酵母菌在醬油醪中的增殖 能力和乙醇醱酵能力也會有很大的差異。 在開放系統進行醱酵及熟化製程所得的普通醬油醪中， 有無數種野生酵母菌共存於其中，並且會形成主要醬油醱 酵酵母菌株。然而，並非所有的主要醬油醱酵酵母菌種在 醬油醪的乙醇醱酵作用中皆擔任同等重要的角色。也就是 說，只有一小部分的醱酵酵母菌種會對乙醇醱酵發揮重要 的作用。 野生酵母菌種可以含有一些具有不全然對醬油品質有利 之性質的微生物。此外，由於其組成會隨著地區或時間而 異，使得產品的品質會因釀造地點的不同而改變，或者是 無法在整年內皆獲得具有一致品質的醬油。 因此，抑制野生酵母菌的成長以維持添加醬油酵母菌的 活潑成長是非常重要的。 依照上述本發明之酵母菌的添加條件，當醬油酵母菌（特 定主要的醬油醱酵酵母菌種）被添加至醬油醪中時，醬油酵 母菌種會與其它菌種競爭以存活，並且它在競爭後不會被 消滅’因此，醬油酵母菌種可以在培養條件下活躍的成長 ’而這些醬油酵母菌種可與野生酵母菌種共存。 接著’依照一般醬油釀造方法，以適當攪伴的方式來控 制醬油乳酸菌和醬油酵母菌所添加進去的醬油醪。接著， 以一般的技術使醬油醪在維持室溫在1 5到35 t (更佳爲20 到 3 0 °C )的條件下進行醱酵和熟成達到3到8個月，以4 -15- 1252737 到6個月爲佳。 將所獲得之成熟醬油醪予以壓榨、過濾，然後予以精製 ，結果就可獲得具有所需乙醇濃度（例如3 % ( V / V )或更高） 、所需乳酸濃度（例如〇 . 9到1 . 〇% )、適當pH値（pH値爲4 . 6 到4 . 9之間）且具有甘醇風味的醬油。 (四）實施方式 實例 以下將以實施例對本發明做更詳細的解釋。 在以下實施例中所使用的醬油酵母菌是藉由已知的一般 方式就可以由醬油醪中輕易地萃取（分離）出來的酵母菌。 這種酵母菌藉由在鹽濃度爲1 6 . 5 ( W / V ) %之醬油麹中進行酒 精醱酵的方式可以生產和累積3 . 0 ( V / V ) %或更高的酒精。 實施例1 將25%(W/V)的鹽水（30升）與依照一般醬油麹菌製法所製 得之醬油麴菌（2 0公斤）摻混在一起，因而製得醬油醪。將 所製得的醬油醪送入一個混合醱酵的容器中。 上述的程序係在使用加熱的蒸氣來消毒製麴菌的裝置以 及混合和醱酵容器的情況下進行，以儘量避免被野生的微 生物（野生酵母菌、乳酸菌等）所污染。 在醬油醪製備好之後’依照一般釀造醬油的方法，使醬 油醪維持在2 0到2 5 °C的溫度下達5個月，並偶爾予以攪 拌，以進行醱酵和熟化。 另一方面’硏究在上述程序中所獲得之醬油醪中的野生 酵母菌隨時間增加和減少的關係，以瞭解混合後下一天的 -16- 1252737 野生酵母菌的數目。 混合後的隔天，將醬油醪予以攪拌，並且加入醬油酵母 菌（純培養溶液）和醬油乳酸菌（純培養溶液）’以達到表1 中所述的條件。此外’在所有的實驗中加入相同數量的乳 酸菌，使得每克醬油醪中所添加的乳酸菌數爲1 X 1 06。在 測試編號6和9的實驗中’加入了給定數量之由上述醬油 醪中分離出的野生酵母菌培養溶液。在其它測試編號的實 驗中，並未添加野生酵母的培養溶液。收集所獲得成熟的 醬油醪液，然後分析每一種液體的組成。此外，也對每一 組液體進行感官測試。結果列於表1。 如表1所示之活菌數測量係依照“食品微生物學手冊”（ 和井久雄，金子安之及山口和夫編撰，技報堂出版，6 0 3 頁）中所述的方法來進行。 如表1所示的醬油成份分析係依照“醬油的實驗方法”（ 日本醬油檢驗所編輯’ 1 9 8 5年3月1日發彳了）一*書中所述 的方法來進行。 12527371252737 发明, the description of the invention (the description of the invention should be described: the technical field, prior art, content, embodiment and schematic description of the invention) (1) Technical field to which the invention belongs: The present invention relates to an improved soy sauce brewing method The method comprises the steps of: mixing soy sauce bacteria into a salt water to make soy sauce mash, adding the pre-cultured soy sauce lactic acid bacteria and soy sauce yeast to the soy sauce mash, performing the fermentation and ripening of the soy sauce mash after adding by a general technique, and A method for obtaining a soy sauce having a desired ethanol concentration and lactic acid concentration and having a glycol flavor. (b) The prior art soy sauce brewing process involves blending steamed soybeans with roasted and milled wheat, inoculating the mixture with seed fungus for soy sauce, and cultivating the mixture. The soy sauce is prepared, and an appropriate amount of salt water is added to the soy sauce to obtain a soy sauce, and after the soy sauce is aged and aged for a fixed period of time, a cooked soy sauce is obtained, and finally the cooked soy sauce is kneaded. Pressure filtration. However, the fermentation and ripening of soy sauce is usually carried out in an open system, and almost no artificial microorganisms such as soy sauce or soy sauce lactic acid bacteria are artificially added. Therefore, all microorganisms (yeast and lactic acid bacteria) which act in soy sauce are natural impurities derived from natural plants inhabiting brewing sites, facilities, devices, etc. In the present invention, 'so-called, wild yeast, , refers to the habitat, facilities, installations, etc. where the soy sauce is brewed, and naturally contaminates the soy sauce and the yeast that proliferates ~ 6 - 1252737. On the contrary, the soy sauce yeast used in the present invention can be easily collected (separated) from soy sauce by a known general method, and is preferably one capable of producing alcohol and being salt-tolerant. This soy sauce yeast is purely cultured and added to the soy sauce. Therefore, this pre-cultured soy sauce yeast is clearly very different from wild yeast. Natural plants are quite diverse, so naturally contaminated soy sauce yeast and soy sauce lactic acid bacteria may contain bacteria that do not have the characteristics of using soy sauce. In addition, due to the difference in the composition of natural plants due to different regions or time, the quality of the products will change with the brewing location, and it will not be able to obtain a certain quality throughout the year. Soy sauce. Based on these viewpoints, the present invention proposes a method of brewing soy sauce in order to continuously produce a brewed soy sauce having a certain quality without considering the limitation of region or time. The method for brewing soy sauce comprises mixing the soy sauce with a salt water to prepare a soy sauce; adding the pre-cultured soy sauce lactic acid bacteria and the soy sauce yeast to the soy sauce; and then performing the soy sauce fermentation after the general technique is added; Ripening. To be more specific, a known method includes mixing soy sauce bacteria into a brine to make soy sauce, adding pre-cultured soy sauce lactic acid bacteria to the soy sauce, and cultivating the bacteria while performing lactate fermentation. (usually 20 to 40 days after mixing), when the number of lactic acid bacteria in the soy sauce is more than 1 〇 7 per gram of soy sauce, and the pH of the soy sauce is reduced to between 5-1 and 5.2. Soy sauce yeast (mainly Zygosaccharomyces rouxii 1252737 'it is the main soy sauce yeast), which is fermented by ethanol, thus producing a good flavor of brewing soy sauce (soy oil science and technology, edited and written by Shinrokuro TOCHIKURA, The Japan Brewing Society, 1 9 8 8 ) (Handbook of Food Microbiology, and Ijiu Hiroshi, edited by Jin Anzhi and Yamaguchi Kazuo, published by Tech Newspaper, 1 9 9 5). However, in this method, since it is not sterile sputum and is fermented and matured in an open container of microorganisms, it cannot be defined based on complicated trouble factors, so lactic acid fermentation and yeast fermentation of soy sauce are very good. It is easily contaminated by wild microorganisms. Therefore, it is difficult to maintain a stable and balanced growth of the added microorganisms for a relatively long period of time, and at the same time, it is necessary to achieve a better fermentation effect. In addition, in the case of pure culture, although the main soy sauce yeast yeast strain has high ethanol productivity or is capable of forming a good soy sauce odor, when these strains are used in a real brewing process, in many cases The same result under pure culture conditions cannot be achieved. Therefore, in recent years, there have been some attempts to achieve balanced and stable long-term growth of the added soy sauce yeast and lactic acid bacteria. To be more specific, even in the latest, sealed and independent mixing and fermentation tanks, and between the adjacent mixing and fermentation tanks, control the fermentation and ripening of the soy sauce, and after each use, Mix the mixing and fermentation tanks or the path of mixing and fermentation. However, because the quality of soy sauce or soy sauce between different batches is also different, pH値 will be unstable with the lactic acid fermentation, which makes this method also have shortcomings. Therefore, it can only be used according to Expert experience -8- 1252737 and the feeling to judge the appropriate time to add yeast. In addition, there is a limit to the complete elimination of microorganisms from the chaotic mixing and fermentation tanks or in the mixing and fermentation pathways. Therefore, when the salt-tolerant wild yeast cannot be eliminated, a small amount of wild yeast remains and contaminates the soy sauce 并且' and may grow into a dominant part before adding the soy sauce yeast, which not only inhibits lactic acid strontium. Yeast, also affected by wild yeast, produces a bad smell. In addition, the added yeast must first be cultured to the required amount and stored. One disadvantage is that when the addition time of the yeast is significantly earlier or later than the predetermined time, it may result in insufficient yeast addition or decreased yeast activity. Therefore, the yeast fermentation is not added to Soy sauce started immediately after the simmering. (3) SUMMARY OF THE INVENTION One of the objects of the present invention is to reliably obtain a desired ethanol concentration (for example, 3% (V / V) or higher), a desired lactic acid concentration (for example, 到. 8 to 1. 〇%),酱 Η値 is a soy sauce having a glycol flavor between 4.6 and 4.9, which solves the above disadvantages by a method of brewing soy sauce, the method comprising mixing the soy sauce bacteria into a brine to prepare In soy sauce, the pre-cultured soy sauce lactic acid bacteria and soy sauce yeast are added to the soy sauce, and after being added by the general technique, the soy sauce is fermented and matured. The focus of the study was on solving the above problems, and the inventors found that by paying attention to the number of cells of the salt-tolerant wild yeast in the soy sauce in the early stage of fermentation, and adding more than the wild yeast in the soy sauce. 1 The soy sauce yeast yeast can achieve the effect of inhibiting the growth of wild yeast, and 9- 1252737 will not affect the lactate fermentation, only activate the growth of the added soy sauce yeast, and can reach Successful ethanol fermentation. In addition, the inventors have also found that when the soy sauce yeast strain is added to the soy sauce in 10 days after mixing, the number of cells of the soy sauce yeast is 10 times higher than that of the wild yeast existing in the soy sauce. By the time of 100 times, it is sufficient to add only a small amount of soy sauce yeast to achieve the above effects. Further, the present inventors have found that during this period, if the addition amount of soy sauce yeast exceeds 2 X 105 cells per gram of soy oleic acid, the fermentation of the lactic acid bacteria coexisting in the soy sauce mash is affected. 'But lactic acid is not produced and accumulated, resulting in poor flavor of soy sauce. Furthermore, the inventors have also found that if the soy sauce yeast is added at a ratio of 2 X 1 〇 5 or less cells per gram of soy sauce, and the number of cells of the soy sauce yeast is higher than that of the wild yeast existing in the soy sauce Ethanol fermentation by 1 〇 to 1 〇〇 曰 ' ' soy sauce yeast can be fully carried out without inhibiting A acid fermentation' and can reliably obtain the desired ethanol concentration (for example 3% ( Soy sauce having a desired lactic acid concentration (for example, 〇. 8 to 〇%), a pH 4 between 4.6 and 4.9, and having a glycol flavor. The present invention has been accomplished by these findings. More specifically, the present invention relates to a method for brewing soy sauce, which comprises the steps of: mixing soy sauce bacteria into a brine to prepare soy sauce; adding pre-cultured soy sauce lactic acid bacteria and soy sauce yeast to the soy sauce. After the addition of the general technique, the fermentation and ripening of the soy sauce is carried out, wherein the k-oil yeast added to the soy sauce is the number of cells of the soy sauce yeast which is larger than the wild yeast which is already present in the soy sauce. Up to 1 times. The invention relates to a method for brewing soy sauce, comprising the steps of: mixing soybean 麴-10- 1252737 bacteria into a brine to prepare soy sauce mash; adding pre-cultured soy sauce lactic acid bacteria and soy sauce yeast to the soy sauce mash; After the addition of the general technique, the soy sauce is fermented and matured, and the soy sauce yeast strain added to the soy sauce is added within 1 day of mixing, and the soy sauce yeast cell number is compared with the wild yeast already present in the soy sauce. In addition, the present invention is a method for brewing soy sauce, comprising the steps of: mixing soy sauce bacteria into a brine to prepare soy sauce; pre-cultured soy sauce lactic acid bacteria and soy sauce The yeast is added to the soy sauce mash; and after being added by the general technique, the soy sauce is fermented and matured, and the soy sauce yeast strain added to the soy sauce mash is added within 10 days of mixing to the soy sauce yeast cell. The number is added 10 to 100 times more than the wild yeast already present in the soy sauce, and is 2 X 105 or less cells per gram of soy sauce. This patent application contains part or all of the contents and/or illustrations of the Japanese Patent Application No. 2001- 387 748, which is the priority file of the present application. The invention will be described in more detail below. In order to carry out the present invention, first, the soy sauce sputum prepared according to the general method for producing soy sauce bacteria is first mixed into salt water according to a general soy sauce brewing method. The amount of brine used herein is one to three times greater than the weight of the soy sauce stock. The salt concentration used herein is adjusted to be suitable for brewing soy sauce, so that the salt concentration is 15 to 20% (w / v), and more preferably 16 to 18% (w / v). When mixing, adjusting the concentration of salt to the above range is the weight of the female, because the salt and the degree of sputum below 15% may cause the soy oleic acid 1252737 to spoil at a temperature of 25 to 35 T: in summer. The lower salt concentration of soy sauce is better for the successful reaction of the enzyme, and it is also beneficial to the proliferation and fermentation of soy sauce lactic acid bacteria and soy sauce yeast, which makes the cells of soy sauce mash and mature more quickly. However, in the case where the salt concentration is less than 15%, the so-called spoilage bacteria will grow in a large amount, thus producing a noticeable acid odor and a rancid taste, which eventually causes the soy sauce product to completely lose its price. Soy sauce 醪 spoilage caused by improper salt concentration can be solved by setting conditions such as maintaining the temperature of soy sauce at 40 ° C or higher, or by adding an acid such as hydrochloric acid or lactic acid. The method adjusts the pH 3 to 3. 〇 or lower. However, these conditions also destroy the original flavor of the soy sauce, thus causing the product to lose the price of soy sauce. When the temperature of the soy sauce or ρί]値 or the nitrogen concentration of the soy sauce is not suitable for the proliferation or survival of spoilage bacteria, the soy sauce will not be spoiled even if the concentration of the salt is low. However, these are only special circumstances and these cannot be considered as a general method of brewing soy sauce. Successful fermentation and ripening cannot be achieved unless the concentration of salt in the summer is 17% or higher or the concentration of salt in the spring or autumn is 15% or higher. In addition, 'hydrolysis of soy sauce at 40 ° C or higher will also lead to poor smell accompanied by the disadvantages of soy sauce, which is called " warm_brew,, (warm) smell. Next 'will A pre-selected or specially cultivated soy sauce lactic acid bacteria and soy sauce yeast (hereinafter sometimes referred to simply as "main soy sauce yeast") are added to the prepared soy sauce, in addition to The soy sauce lactic acid bacteria can be added in accordance with a conventional method of brewing soy sauce in a ratio of 1 X 4 cells per gram of soy sauce to 1 X 1 07 cells per gram of soy sauce. - 1 2 - 1252737 It is quite important in the present invention that the soy sauce yeast added allows the soy sauce yeast to have a cell number of 10 to 1000 times more than the wild yeast which is already present in the soy sauce. When this ratio is lower than 0 times, there is a danger that the wild yeast bacteria coexisting in the soy sauce mash becomes a relatively large number, and the soy sauce yeast strain which has good properties and has been added to the soy sauce mash is difficult to grow. Re-growth '迨 is due to its competition with wild yeast coexisting in soy oleic acid, so these soy sauce yeasts may be destroyed or expelled under competitive growth inhibition. Conversely, 'if the ratio of addition is more than 100 times, the coexisting lactic acid bacteria cannot grow vigorously', so the desired lactic acid concentration cannot be obtained, and the appropriate enthalpy (for example, ρ Η値 is between 4.6 and 4.9). ) and a brewed soy sauce with a flavor of glycol. Further, the pre-cultured soy sauce lactic acid bacteria and soy sauce yeast are preferably added within 1 day after mixing, and it is preferably added within seven days after mixing. The best addition time is between the time of mixing (just after mixing) and the next day after mixing. The reason this range is more appropriate is because it can significantly reduce the amount of soy sauce added. Conversely, when the soy sauce yeast is added in the day after mixing or later, the number of wild yeasts coexisting in the soy sauce mash becomes more. In this case, it is necessary to add a larger amount of soy sauce yeast. As a result, when the concentration of the _ mother bacteria becomes higher, the growth of the lactic acid bacteria is suppressed, so that such a result will cause the obtained soy sauce to have a low lactic acid content and the wind is not bad. In addition, there is a danger that the stomach 'oil yeast (the main soy sauce yeast with good properties) added to the soy sauce oysters is difficult to live - 13 - 1252737 splash growth 'this is due to its coexistence with The competition for wild yeast in soy sauce ''s therefore 'under the competitive growth inhibition of wild yeast', these soy sauce yeasts may have been eliminated or expelled. That is to say, since the soy sauce yeast may not be effectively proliferated, the problem of the present invention cannot be solved smoothly. Quite importantly, the amount of soy sauce added is 2 X 1 05 or less per cell per gram of soy sauce. When the ratio is 2 X 105 cells per gram of soy sauce or higher, the lactic acid bacteria will not be effectively swelled. Therefore, it may be impossible to obtain a soy sauce having a desired lactic acid concentration, an appropriate p pj 値 (for example, ρ η 値 between 4.6 and 4.9), and a glycol flavor. Μ Examples of the soy sauce yeast of the present invention include a salt-tolerant soy sauce yeast yeast for brewing soy sauce, such as: Z yg 〇sacchar 〇mycesr 〇u X ii strain ' and is mainly suitable for ethanol fermentation of soy sauce 醪The main soy sauce yeast yeast 'in the presence of 6 to 28% (w / v) of sodium chloride and glucose 'has: has considerable ethanol fermentation capacity; belongs to the Candida genus 'includes' Candida versatil is and Candida etch ellsii's salt-tolerant yeast scorpion scorpion; or the salt-tolerant yeast group of the genus of the genus of the genus of the genus of the genus of the genus of the genus of the genus of the genus of the genus Mainly related to the general aroma of soy sauce, sometimes referred to as the late ripe yeast group. The main soy sauce having the ability to carry out active ethanol fermentation in the yeast group is preferably obtained by (especially) having an ethanol concentration of 3 % (V / V) or higher and a flavor of glycol. According to the inventors' research, the main soy sauce yeasts cannot be regarded as common to each other because they vary depending on the species. That is to say, the proliferative ability of the main soy sauce yeast of different strains in soy sauce and the ability to eluate ethanol can be greatly different. Among the common soy sauces obtained by the open system for the fermentation and ripening process, there are a myriad of wild yeasts coexisting therein, and a main soy yeast yeast strain is formed. However, not all of the main soy yeast yeasts play an equally important role in the ethanol fermentation of soy sauce. That is to say, only a small portion of the yeast yeast species play an important role in ethanol fermentation. Wild yeast species may contain microorganisms that have properties that are not entirely beneficial to the quality of the soy sauce. In addition, since the composition varies depending on the region or time, the quality of the product may vary depending on the brewing location, or it may not be possible to obtain a consistent quality of soy sauce throughout the year. Therefore, it is very important to inhibit the growth of wild yeast to maintain the viable growth of soy sauce yeast. According to the above-described conditions for the addition of the yeast of the present invention, when soy sauce yeast (specific major soy sauce yeast yeast species) is added to the soy sauce, the soy sauce yeast species will compete with other strains to survive, and it is After competition, it will not be eliminated. Therefore, soy sauce yeast species can grow actively under culture conditions, and these soy sauce yeast species can coexist with wild yeast species. Then, according to the general soy sauce brewing method, the soy sauce mash added by the soy sauce lactic acid bacteria and the soy sauce yeast is controlled in an appropriate manner. Next, the soy sauce is fermented and matured at a temperature of 15 to 35 t (more preferably 20 to 30 ° C) for 3 to 8 months by a general technique, to 4 -15- 1252737 to 6 months is preferred. The obtained soy sauce mash is pressed, filtered, and then refined, and as a result, a desired ethanol concentration (for example, 3% (V / V) or higher) and a desired lactic acid concentration (for example, 〇. 9 to 1) can be obtained. 〇%), a suitable pH 値 (pH 4 between 4.6 and 4.9) and a soy sauce with a sweet flavor. (4) Embodiments The present invention will be explained in more detail by way of examples. The soy sauce yeast used in the following examples is a yeast which can be easily extracted (separated) from soy sauce by a known general method. This yeast can produce and accumulate 3.0 (V / V ) % or higher alcohol by alcohol fermentation in a soy sauce with a salt concentration of 16.5 (W / V ) %. Example 1 25% (w/v) of saline (30 liters) was blended with soy sauce (20 kg) prepared according to the general soy sauce method, thereby preparing soy sauce. The prepared soy sauce is poured into a mixed fermented container. The above procedure is carried out in the case of using a heated vapor to sterilize the sputum-producing device and mixing and fermenting the container to avoid contamination by wild microorganisms (wild yeast, lactic acid bacteria, etc.). After the soy sauce is prepared, the soy sauce is maintained at a temperature of 20 to 25 ° C for 5 months in accordance with a general method of brewing soy sauce, and occasionally stirred for fermentation and ripening. On the other hand, the relationship between the increase and decrease of wild yeast in the soy sauce mash obtained in the above procedure was observed to understand the number of -16-1252737 wild yeasts after the next day of mixing. The next day after mixing, the soy sauce was stirred, and soy sauce yeast (pure culture solution) and soy sauce lactic acid bacteria (pure culture solution) were added to achieve the conditions described in Table 1. In addition, the same amount of lactic acid bacteria was added to all experiments so that the number of lactic acid bacteria added per gram of soy sauce was 1 X 1 06. In the experiments of Test Nos. 6 and 9, 'a given amount of the wild yeast culture solution separated from the above soy sauce was added'. In the experiment of other test numbers, the culture solution of wild yeast was not added. The mature soy sauce mash was collected and analyzed for the composition of each liquid. In addition, sensory tests were performed on each set of liquids. The results are shown in Table 1. The measurement of the viable count as shown in Table 1 was carried out in accordance with the method described in the "Handbook of Food Microbiology" (and Ichio Hiroshi, edited by Jin Anzhi and Yamaguchi Kazuo, Technical Newspaper, pp. 603). The analysis of the soy sauce composition shown in Table 1 was carried out in accordance with the method described in the "Experimental Method of Soy Sauce" (edited by the Japanese Soy Sauce Inspection Institute on March 1, 1985). 1252737
A 1 c代表乙醇，L a c代表乳酸 “氣味”和“味覺”欄中的符號： 良好，△代表有點差，X代表不良 -- _PH 氣味 味覺 品評 4.77 Δ 〇 酸味 4.85 ◎ ◎ 風味 良好 4.86 ◎ ◎ 風味 良好 4.86 ◎ ◎ 風味 良好 4.88 ◎ ◎ 風味 良好 4,89 ◎ 〇 風味 良好 5.10 〇 Δ 混濁 不淸 5.28 〇 X 殘餘 風味 5,08 〇 Δ 混濁 不淸 ◎代表非常好，〇代表 由表1中的結果可淸楚的得到以下幾點事實： ⑴在添加醬油酵母囷倍率爲5倍的比較實例i之測試結 果中可淸楚的知道’所得之成熟醬油醪具有酸味，而無法 獲得具有芳醇香味的醬油。說的更明確一點，當醬油酵母 菌的添加倍率小於1 〇倍時’在醬油酸中所共同存在的野生 酵母菌就變成相對多數’因此，具有良好性質的醬油酵母 菌就很難存活’在與共存於醬油醪中之野生酵母菌的競爭 ~18~ 1252737 性抑制生長的結果之下’這些醬油酵母菌截就被消滅了。 (2 )在添加醬油酵母菌倍率分別爲2〇〇倍和4〇〇倍的比較 實例2和3之测試結果中可淸楚的知道，乳酸濃度明顯偏 低。因此’無法獲得具有獨特甘醇風味的请油。 (3 )即使在添加醬油酵母菌倍率爲1 Q 〇倍的比較實例4之 測§式結果中仍可淸楚的知道，當每克醬油醪中含有酵母菌 數超過2 X 1 05時，乳酸濃度明顯偏低。結果將無法獲得具 有所需乳酸濃度、適當pH値（例如PH値爲4 . 6到4 . 9之間） 以及甘醇風味的醬油。 (4 )與上述這些結果相反的是’在本發明實例的五項測試 中’其中醬油酵母菌細胞數爲其它的1 〇到丨〇 〇倍，且添加 之醬油酵母菌的比率爲每克醬油醪中含有2 X丨〇5或更少個 細胞數’藉由添加醬油酵母菌可以劇烈的進行乙醇醱酵， 而可獲得具有3.0%(V/V)或更高，或者特別是高達3.5(v/V) 或更高之目標乙醇濃度的醬油。此外，很清楚的是，較好 是以醬油乳酸菌來進行乳酸醱酵，而可獲得具有適當乳酸 含量的醬油，其P Η値爲4 . 8到4 · 9之間。因此，整體而言 ’本發明可以得具有特殊香味及良好風味的醬油。 實施例2 製備8項測試所需的2 0公斤醬油麴菌，每一個皆是依照 一般醬油麴菌製法所製得。將醬油麴菌和2 5 % ( w / ν )的鹽水 (3 0升）分別添加至8個醱酵容器中，然後混合製得醬油醪 。一旦混合之後，將分離自醬油||的醬油乳酸菌以1 X 1 〇6 個細胞/克醬油醪的比例添加進去，並且野生酵母菌的添加 - 1 9 - 1252737 比率爲2 X 1 〇3個細胞/克醬油醪，接著依照傳統釀造 的方法來控制醬油麴，以進行醱酵和熟化。 另一方面’硏究在上述程序中所獲得之醬油醪中的 酵母菌隨時間增加和減少的關係，以瞭解混合後 1 , 3 , 5，7 , 1 〇，1 2和2 0天的野生酵母菌的數目。 接著，除了第1項測試之外，將表2中所述的醬油 菌分別在第丨天添加至第二項測試、第3天添加至第 測試、第5天添加至第四項測試、第7天添加至第五 試第1 0天添加至第六項測試、第1 2天添加至第七項 ，及第20天添加至第八項測試中以進行後醱酵作用。 及醱酵的容器係置於一個溫度爲2 3 °C的溫控室中’然 照傳統釀造醬油的方法使這些醬油醪釀造五個月’以 醱酵和熟化。 收集所製得的醬油醪液，然後施以類似實施例1中 的分析方法及感官品評。結果列於表2。 醬油 野生 的第 酵母 三項 項測 測試 混合 後依 進行 所述 -20- I252737 袠2 _式 欏號 ^—__ 項目 添加之培 養酵母菌 (天） 耐鹽野生酵 母菌(A) 經培養的酵 母菌(B) B/A (倍率） 成份分析 氣味 味覺 品評 Ale Lac pH 1 本發明 〇 2X103 2X 104 10 3.63 0.92 4.85 ◎ ◎ 風味 ----_ 良好 2 本發明 1 2X103 3x 104 15 3.50 0.90 4.88 ◎ ◎ 風味 〜--- 良好 3 本發明 3 5X103 6X 104 12 3.55 0.87 4.88 ◎ ◎ 風味 ----- 良好 4 本發明 5 7X103 8X 104 11 3.60 0.91 4.85 ◎ ◎ 風味 --- 良好 5 本發明 7 8X103 1X105 13 3.67 0.90 4.90 ◎ ◎ 風味 良好 6 本發明 10 1.2X104 1.5X105 13 3.65 0.88 4.90 〇 〇 風味 良好 7 比較實例1 12 2.2X104 3X105 13 3.65 0.15 5.12 〇 Δ 混濁 不淸 8 比較實例2 20 1.3X105 2X106 15 3.63 0.09 5.22 〇 X 殘餘 風味 由表2的結果可得知，醬油酵母菌較好是在混合後1 〇天 以內添加。當醬油酵母菌在混合時添加或者是在混合後七 天內添加時，很明顯地，可獲得具有所需乙醇及乳酸濃度 且具有甘醇風味的醬油。 此外，當醬油酵母菌的添加時間在混合1 〇天之後時，共 存在醬油醪中的野生酵母菌數變得比其它菌數多得多，医! 此無可避免的要添加更大量的醬油酵母菌。由於酵母菌Μ 度的增加會抑制乳酸菌的生長，因此很明顯地的，這樣白勺 結果將會造成所獲得的醬油具有低的乳酸含量且風味；^ ^ 實例3 -21- 1252737 依照類似實施例1的方式來製備醬油醪。在混合之後的 那一天，將醬油醪予以攪拌，然後以丨X丨〇3個細胞/克醬 油酸的比例來添加野生酵母菌，以及以1 X 1〇6個細胞/克 醬油醪的比率來添加乳酸菌。在本發明方法中，當添加野 生酵母菌時，另以1 X 1 0 4個細胞/克醬油醪的比率來添加 良好的培養乳酸菌，使得醬油酵母菌的細胞數比野生酵母 菌數大1 0倍。在對照組的實驗方法中，當醬油醪的p[1値 變爲5 . 2或更低時，以2 X 1 〇5個細胞/克醬油醪的比率來 添加良好的培養乳酸菌。每一組添加皆進行五次。表3所 示爲醬油醪液在混合1 3 5天之後的乳酸含量（w / V% )。此外 醬油醪的溫度與實施例1相同。 表3 乳酸（W / V ) % 實驗編號 對照組 本發明 1 1.00 0.95 2 0.91 0.93 3 0.98 0.96 4 0.72 0.95 5 1.06 0.91 平均値 0.93 0.94 標準値 _ 0.13 0.02 ® ^ 3的結果可淸楚的看出：由於在對照組實驗中的乳 酉妾t胃'變化極大，因此很難獲得具有均勻乳酸含量的醬油 〇 相反地’由於本發明方法中的乳酸含量幾乎相同，因此 可以獲得具有均勻乳酸含量的醬油。 -22- 1252737 本發明效果 依照本發明，添加至醬油醪中的醬油酵母菌的活菌細胞 數比既存於醬油醪中的野生酵母菌多1 0倍到1 〇〇倍，因此 可以成功的進行酵母菌醱酵，而不會影響乳酸醱酵，其同 時可抑制野生酵母囷的成長’並且只會使添加的醬油酵母 菌活潑的成長。 此外’依照本發明，醬油酵母菌係在混合後的1 0天之內 添加至醬油醪中，而使得醬油酵母菌的細胞數比既存於醬 油麹中的野生醱母菌多1 0倍到1 00倍。因此，上述的效果 可藉由添加極少量的醬油酵母菌而由本發明達成。 此外’在此同時’依照本發明，在混合後丨0天內添加至 醬油醪中的醬油酵母菌細胞數比既存於醬油醪中的野生酵 母菌多1 0倍到1 0 0倍’並且相對於醬油麴而言，每克醬油 _中含有2 X 1 05或更少個細胞。因此，醬油酵母菌可充分 的進行乙醇醱酵作用，而不會抑制乳醱酵，所以可以可靠 地製得具有所需乙醇濃度（例如3 % ( V / v )或更高）、所需乳 酸濃度（例如〇 · 8到1 · 〇%)、pH値爲4 . 6到4 . 9之間且具有 甘醇風味的醬油。A 1 c stands for ethanol, and L ac stands for the symbols in the "smell" and "taste" columns of lactic acid: good, △ stands for a little bad, X stands for bad - _PH odor taste tastes 4.77 Δ 〇 sour taste 4.85 ◎ ◎ good flavor 4.86 ◎ ◎ Good flavor 4.86 ◎ ◎ Good flavor 4.88 ◎ ◎ Good flavor 4,89 ◎ Good flavor 5.10 〇Δ turbidity 5.28 〇X Residual flavor 5,08 〇Δ turbidity ◎ representative is very good, 〇 represents by Table 1 As a result, the following facts can be obtained: (1) It is succinct to know that the obtained mature soy sauce has a sour taste and cannot obtain an aromatic alcohol flavor in the test result of the comparative example i in which the soy sauce yeast has a magnification of 5 times. soy sauce. To be more specific, when the addition rate of soy sauce yeast is less than 1 〇 times, the wild yeast that coexists in soy oleic acid becomes relatively large. Therefore, soy yeast with good properties is difficult to survive. Competing with the wild yeast coexisting in soy sauce ~~18~ 1252737 The results of the inhibition of growth were 'these soy sauce yeast cuts were eliminated. (2) In comparison with the test results of Examples 2 and 3, the ratios of the soy sauce yeasts were 2 〇〇 and 4 分别, respectively, and it was found that the lactic acid concentration was significantly lower. Therefore, it is not possible to obtain oil with a unique glycol flavor. (3) Even in the results of the comparative example 4 of the comparative example 4 in which the soy sauce yeast ratio is 1 Q 添加, it is still known that lactic acid is contained when the number of yeasts per gram of soy sauce contains more than 2×10 5 The concentration is significantly lower. As a result, it is impossible to obtain a soy sauce having a desired lactic acid concentration, an appropriate pH (for example, a pH of 4.6 to 4.9), and a glycol flavor. (4) Contrary to the above results, 'in the five tests of the examples of the present invention, the number of cells of the soy sauce yeast is 1 〇 to 丨〇〇 times, and the ratio of the added soy sauce yeast is per gram of soy sauce. The sputum contains 2 X 丨〇 5 or less cells. 'According to the soy sauce yeast, the ethanol can be fermented violently, and it can be obtained with 3.0% (V/V) or higher, or especially up to 3.5 ( Soy sauce with a target ethanol concentration of v/V) or higher. Further, it is clear that it is preferred to carry out lactic acid fermentation with soy sauce lactic acid bacteria to obtain a soy sauce having an appropriate lactic acid content, and the P Η値 is between 4.8 and 4.9. Therefore, as a whole, the present invention can provide a soy sauce having a special aroma and a good flavor. Example 2 20 kg of soy sauce bacteria required for the eight tests were prepared, each of which was prepared according to the general soy sauce method. Soy sauce soy and 25% (w / ν) of brine (30 liters) were separately added to 8 fermented containers, and then mixed to prepare soy sauce 醪. Once mixed, the soy sauce lactic acid bacteria isolated from soy sauce|| was added in a ratio of 1 X 1 〇 6 cells/g soy sauce, and the addition of wild yeast was -1 9 - 1252737 in a ratio of 2 X 1 〇 3 cells. / g soy sauce, then control the soy sauce according to the traditional brewing method for fermentation and ripening. On the other hand, 'study on the relationship between the increase and decrease of yeast in soy sauce obtained in the above procedure over time to understand the wild after mixing 1, 3, 5, 7, 1 , 12 and 20 days. The number of yeasts. Next, in addition to the first test, the soy sauces described in Table 2 were added to the second test on the third day, the third test was added on the third day, and the fourth test was added on the fifth day. 7 days added to the fifth test, the 10th day was added to the sixth test, the 12th day was added to the seventh item, and the 20th day was added to the eighth test for post-fermentation. The fermented containers were placed in a temperature-controlled room at a temperature of 23 ° C. These sauces were brewed for five months by means of traditional brewing soy sauce to ferment and ripen. The prepared soy sauce mash was collected, and then subjected to an analysis method similar to that in Example 1 and a sensory evaluation. The results are shown in Table 2. The soy sauce wild yeast three test tests are mixed after the -20- I252737 袠2 _ 椤 ^ ^___ project added culture yeast (days) salt-tolerant wild yeast (A) cultured yeast Bacteria (B) B/A (magnification) Ingredient analysis Odour taste evaluation Ale Lac pH 1 The present invention 〇 2X103 2X 104 10 3.63 0.92 4.85 ◎ ◎ Flavor----_ Good 2 The present invention 1 2X103 3x 104 15 3.50 0.90 4.88 ◎ ◎ flavor ~--- good 3 invention 3 5X103 6X 104 12 3.55 0.87 4.88 ◎ ◎ flavor ----- good 4 invention 5 7X103 8X 104 11 3.60 0.91 4.85 ◎ ◎ flavor --- good 5 invention 7 8X103 1X105 13 3.67 0.90 4.90 ◎ ◎ Good flavor 6 Inventive 10 1.2X104 1.5X105 13 3.65 0.88 4.90 Good flavor 7 Comparative Example 1 12 2.2X104 3X105 13 3.65 0.15 5.12 〇Δ turbidity 8 Comparative example 2 20 1.3X105 2X106 15 3.63 0.09 5.22 〇X Residual flavor can be known from the results in Table 2, soy sauce yeast is better mixed Within days after adding 1 billion. When soy sauce yeast is added during mixing or added within seven days after mixing, it is apparent that soy sauce having a desired ethanol and lactic acid concentration and having a glycoly flavor can be obtained. In addition, when the addition time of soy sauce yeast is mixed after 1 day, the number of wild yeasts coexisting in soy sauce mash becomes much more than other bacteria, so it is inevitable to add a larger amount of soy sauce. Yeast. Since the increase in yeast temperature inhibits the growth of lactic acid bacteria, it is apparent that such a result would result in a soy sauce having a low lactic acid content and flavor; ^ ^ Example 3 - 21 - 1252737 in accordance with a similar example 1 way to prepare soy sauce 醪. On the day after the mixing, the soy sauce was stirred, and then the wild yeast was added at a ratio of 丨X丨〇3 cells/g soy sauce, and at a ratio of 1 X 1 6 cells/g soy sauce. Add lactic acid bacteria. In the method of the present invention, when wild yeast is added, a good cultured lactic acid bacterium is added at a ratio of 1×10 4 cells/g soy sauce, so that the number of cells of the soy sauce yeast is larger than that of the wild yeast. Times. In the experimental method of the control group, when p[1値 of soy sauce was changed to 5.2 or lower, a well-cultured lactic acid bacterium was added at a ratio of 2 X 1 〇 5 cells/g soy sauce. Each set is added five times. Table 3 shows the lactic acid content (w / V%) of the soy sauce mash after mixing for 135 days. Further, the temperature of the soy sauce was the same as in Example 1. Table 3 Lactic acid (W / V) % Experimental number control group The present invention 1 1.00 0.95 2 0.91 0.93 3 0.98 0.96 4 0.72 0.95 5 1.06 0.91 The average 値 0.93 0.94 standard 値 _ 0.13 0.02 ® ^ 3 results can be clearly seen : Since the chyle t stomach 'variation in the control experiment is extremely large, it is difficult to obtain a soy sauce having a uniform lactic acid content. Conversely, since the lactic acid content in the method of the present invention is almost the same, it is possible to obtain a uniform lactic acid content. soy sauce. -22- 1252737 EFFECT OF THE INVENTION According to the present invention, the number of viable cells of soy sauce yeast added to soy sauce is 10 to 1 times higher than that of wild yeast existing in soy sauce, so that it can be successfully carried out The yeast is fermented without affecting the lactic acid fermentation, which at the same time inhibits the growth of wild yeast cockroaches' and only makes the added soy sauce yeast grow vigorously. In addition, according to the present invention, the soy sauce yeast strain is added to the soy sauce mash within 10 days after mixing, so that the number of cells of the soy sauce yeast is 10 times to 1 more than that of the wild sputum bacteria existing in the soy sauce mash. 00 times. Therefore, the above effects can be achieved by the present invention by adding a very small amount of soy sauce yeast. Further, 'at the same time', according to the present invention, the number of soy sauce yeast cells added to the soy sauce mash within 0 days after mixing is 10 times to 100 times more than that of the wild yeast existing in the soy sauce mash and relative In soy sauce, there are 2 X 105 or fewer cells per gram of soy sauce. Therefore, soy sauce yeast can fully carry out ethanol fermentation without inhibiting lactolysis, so it is possible to reliably produce a desired lactic acid having a desired ethanol concentration (for example, 3% (v / v) or higher). Soy sauce having a concentration of (for example, 〇·8 to 1 · 〇%) and having a pH of between 4.6 and 4.9 and having a glycol flavor.
Ifc外'’即丨吏医1爲不同批而造成醬油麴菌或醬油醪不同時 ’本發明仍可獲得所需之可靠醬油。 Λ有*在經過混合後不久的幾天內（特別是在混合後的i 〇 X t Θ )添加醬油酵母菌，並且測量已存在醬油醪中之野生 _ m ® % ^目’才能夠決定具有最佳乙醇醱酵能力之醬油 B母：® @ ®當添加量。除此之外，也不需擔心添加醬油酵 -23- 1252737 母菌的時間被延遲，因而可以依照所需規劃來準備醬油酵 母菌的純培養產物的適當數量。 此外，倚賴專家經驗和感覺所做的判斷就變得不需要了 〇 此外，即使因爲受限於無法自淸潔的醱酵槽和醱酵路徑 中完全淸除微生物而無法消滅耐鹽性的野生酵母菌，導致 有少數的野生酵母菌殘留並且污染了醬油醪，但以高於不 要之酵母菌種數倍之固定比率來添加醬油酵母菌可以有效 的抑制野生酵母菌增殖，並且可以只促進醬油酵母菌之活 化增生。 此外，酵母菌的添加量皆維持在不會抑制乳酸醱酵作_ 的範圍內，因此可使得乳酸醱酵得以成功的進行，並且製 得具有適當乳酸含量的醬油。 因此，將本發明應用於一般已知的醬油釀造法中，可以 獲得具有所需乙醇和乳酸濃度且具甘醇風味的醬油，例如 濃口醬油、薄口醬油、白醬油、壺底醬油、再醱酵醬油或 醬油膏。 所有在本文中所述引用的刊物、專利及專利申請書皆全 部倂爲本申請書之參考內容。 圖式簡單說明：無 -24-Ifc is ''that is, when the doctor 1 is different batches and the soy sauce or soy sauce is different, the present invention still obtains the desired reliable soy sauce. Λ有* In the few days after mixing (especially after mixing i 〇X t Θ ) add soy sauce yeast, and measure the presence of wild _ m ® % ^ in the soy sauce 才 can be decided to have The best ethanol fermentation capacity of soy sauce B mother: ® @ ® when added. In addition, there is no need to worry about the delay in adding the soy sauce -23- 1252737 mother bacterium, so that the appropriate amount of pure culture product of the soy sauce yeast can be prepared according to the required plan. In addition, judgments based on expert experience and feelings become unnecessary. In addition, even if it is limited by the inability to cleanse the fermenting tank and the fermentation path to completely eliminate microorganisms, it is impossible to eliminate the salt-tolerant wild. Yeast, which causes a small amount of wild yeast to remain and contaminates the soy sauce, but the addition of soy sauce yeast at a fixed ratio several times higher than the undesired yeast species can effectively inhibit the proliferation of wild yeast and can only promote soy sauce. Activated proliferation of yeast. Further, the amount of the yeast added is maintained within a range that does not inhibit the lactic acid fermentation, so that the lactic acid fermentation can be successfully carried out, and the soy sauce having an appropriate lactic acid content can be obtained. Therefore, by applying the present invention to a generally known soy sauce brewing method, it is possible to obtain a soy sauce having a desired ethanol and lactic acid concentration and having a glycol flavor, such as concentrated soy sauce, thin mouth soy sauce, white soy sauce, pot bottom soy sauce, and further fermentation. Soy sauce or soy sauce. All publications, patents, and patent applications cited in this specification are hereby incorporated by reference. Simple description: no -24-